1. Field of the Invention
This application relates to human H37 protein and to cDNA encoding the protein. More particularly, this application relates to human H37 protein that is an activity-controlling subunit for the protein Cdc7 controlling the replication of human cells; to human gene encoding the protein; to an antibody to the H37 protein; and to a method for controlling the proliferation of human cells using such genetic engineering material and antibody.
2. Description of the Related Art
Proliferation of cells is initiated when a liquid factor called a growth factor is bound to the receptor on the cell surface and a signal for proliferation is transmitted into the cell. Accordingly, for an artificial induction of proliferation of incubated cells, methods where an excessive amount of a growth factor is added to a cell medium, where a receptor which is not inherently owned by the said cell is expressed on the cell surface and a factor which is specific for the receptor is added to a medium, etc. have been carried out. Further, in suppressing the cell proliferation, methods where competing molecule, antagonist or the like to the receptor protein is added to the medium to suppress the binding of the receptor to the growth factor, etc. have been carried out.
On the other hand, in the case of the cell where a proliferation signal is issued by binding of the receptor to the growth factor, a cycle in which its genomic DNA is replicated, uniformly distributed to daughter cells and then divided is repeated. Such a cycle is called “cell cycle” especially for eukaryotes. The cell cycle is basically classified into four phases. Thus, S phase when chromosomal DNA is replicated; M phase when the replicated chromosome is divided by a spindle body and then cytoplasm is divided; G1 phase which is a period from M phase finishes until S phase begins; and G2 phase which is a period from S phase finishes until M phase starts. Transition from G1 phase to S phase is particularly strictly controlled and DNA replication takes place only once in S phase.
It has been confirmed from the studies in yeast and higher eukaryotic cells that cyclin-dependent kinase plays a critical role in the cell cycle progression (Nature 292:558-560, 1981; Cell 66:731-742, 1991; Nature 349:338393, 1991; Science 257:1958-1961, 1992; Bioassays 17:471, 1995). Further, from a genetic analysis in yeast, it has been clarified that another serine/threonine linase plays an essential role in the initial stage of S phase (G1/S transition). Thus, characterization of cdc7 mutation which was isolated as one of the cell division cycle mutants (J. Mol. Biol. 59:183-194, 1971), it revealed that the Cdc7 protein kinase functions immediately prior to chromosomal replication and that, during S phase, that is necessary for activation of origins (Mol. Cell Biol. 6:1590-1598, 1986; Genes Dev. 15:480-490, 1998; Genes Dev. 15:491-501, 1998). It has been also clarified that the Cdc7 kinase activity is dependent upon the presence of a regulatory subunit, Dbf4 (Genetics 131:21-29, 1992; Mol. Cell. Biol. 13:2899-2908, 1993). Expression of Dbf4 is periodic and is regulated by both at the transcriptional and post-translational levels (Exp. Cell Res. 180:419-428, 1989). The increase in Cdc7 kinase activity at the G1/S boundary is at least accounted for by the elevated expression of Dbf4 in late G1 phase (Mol. Cell. Biol. 13:2899-2908, 1994; Exp. Cell Res. 180:419-428, 1989). In addition, since Dbf4 interacts with replication origins in vivo (Science 265:1243-1246, 1994), it has been suggested that the Cdc7 may trigger S phase by directly activating the replication initiation complex assembled at the origins.
In addition, the inventors of this application had already isolated kinases related to yeast Cdc7 from Schizosacchatromyces prombe, Xenopus, mouse and human (J. Biol. Chem. 273:23248-23257, 1998; EMBO J. 16:4340-4351, 1997; EMBO J. 14:3094-3104, 1995), and pointed out that eukaryotic chromosomal replication is regulated by a conserved mechanism involving this family of kinase.
From the findings in yeast and higher eukaryotes as mentioned above, it is expected that an artificial control of the cell proliferation is possible by means of the regulation of the Cdc7 kind activity in cells, which is entirely other means than conventional methods using operation of growth factor/receptor binding.
However, the inventors had also found that the putative human homolog of Cdc7, huCdc7, possesses only a very low level of kinase activity when overexpressed in mammalian cells while a baculovirus expressed form of huCdc7 is inactive.
Under such circumstances, the inventors of this application have investigated a human cDNA library with a presumption of the presence of a regulatory subunit for human Cdc7, succeeded in isolating the cDNA encoding a novel protein which regulates the kinase activity by binding to huCdc7 and named the said protein encoded in this cDNA as H37 protein.
An object of the invention is to provide the novel protein obtained by the inventors in an industrial applicable form.
Another object of the invention is to provide a human gene encoding the protein and to provide a material for genetic operation including cDNA derived from the gene, an antibody to the protein.
Still another object of the invention is to provide a method for the artificial control of proliferation of human cells using the above material for genetic operation.